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An essential regulatory function of the DnaK chaperone dictates the decision between proliferation and maintenance in Caulobacter crescentus
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab). Philipps University Marburg, Germany.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute. Stockholm University, Science for Life Laboratory (SciLifeLab). Philipps University Marburg, Germany.ORCID iD: 0000-0001-9150-3217
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Number of Authors: 52017 (English)In: PLoS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 13, no 12, article id e1007148Article in journal (Refereed) Published
Abstract [en]

Hsp70 chaperones are well known for their important functions in maintaining protein homeostasis during thermal stress conditions. In many bacteria the Hsp70 homolog DnaK is also required for growth in the absence of stress. The molecular reasons underlying Hsp70 essentiality remain in most cases unclear. Here, we demonstrate that DnaK is essential in the alpha-proteobacterium Caulobacter crescentus due to its regulatory function in gene expression. Using a suppressor screen we identified mutations that allow growth in the absence of DnaK. All mutations reduced the activity of the heat shock sigma factor sigma(32) , demonstrating that the DnaK-dependent inactivation of sigma(32) is a growth requirement. While most mutations occurred in the rpoH gene encoding sigma(32) , we also identified mutations affecting sigma(32) activity or stability in trans, providing important new insight into the regulatory mechanisms controlling sigma(32) activity. Most notably, we describe a mutation in the ATP dependent protease HslUV that induces rapid degradation of sigma(32) , and a mutation leading to increased levels of the house keeping sigma(70) that outcompete sigma(32) for binding to the RNA polymerase. We demonstrate that sigma(32) inhibits growth and that its unrestrained activity leads to an extensive reprogramming of global gene expression, resulting in upregulation of repair and maintenance functions and downregulation of the growth-promoting functions of protein translation, DNA replication and certain metabolic processes. While this re-allocation from proliferative to maintenance functions could provide an advantage during heat stress, it leads to growth defects under favorable conditions. We conclude that Caulobacter has coopted the DnaK chaperone system as an essential regulator of gene expression under conditions when its folding activity is dispensable.

Place, publisher, year, edition, pages
2017. Vol. 13, no 12, article id e1007148
National Category
Biological Sciences
Research subject
Molecular Bioscience
Identifiers
URN: urn:nbn:se:su:diva-152516DOI: 10.1371/journal.pgen.1007148ISI: 000419103000028PubMedID: 29281627OAI: oai:DiVA.org:su-152516DiVA, id: diva2:1180927
Available from: 2018-02-07 Created: 2018-02-07 Last updated: 2019-02-12Bibliographically approved
In thesis
1. Coping with Stress: Regulation of the Caulobacter crescentus cell cycle in response to environmental cues
Open this publication in new window or tab >>Coping with Stress: Regulation of the Caulobacter crescentus cell cycle in response to environmental cues
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

All organisms have to respond to environmental changes to maintain cellular and genome integrity. In particular, unicellular organisms like bacteria must be able to analyze their surroundings and rapidly adjust their growth mode and cell cycle program in response to environmental changes, such as changes in nutrient availability, temperature, osmolarity, or pH. Additionally, they have to compete with other species for nutrients and evade possible predators or the immune system. Bacteria exhibit a myriad of sophisticated regulatory pathways that allow them to cope with various kinds of threats and ensure their survival. However, the precise molecular mechanisms underlying these responses remain in many cases incompletely described. This thesis focuses on the mechanisms that adjust growth and cell cycle progression of Caulobacter crescentus under adverse conditions.

In paper I we describe a mechanism by which environmental information is transduced via the membrane-bound cell cycle kinase CckA into the cell division program of C. crescentus. This mechanism ensures rapid dephosphorylation and clearance of the cell cycle master regulator CtrA under salt and ethanol stress. The downregulation of CtrA leads to a cell division block and cell filamentation, which provides a growth advantage under these conditions.

Cell filamentation of C. crescentus can also be observed in the late stationary phase, in which a small subpopulation of cells transforms into helical shaped filaments. In these cells not only CtrA but all major cell cycle regulators are cleared (paper II), leading to a situation in which cells block their cell cycle but continue to grow. We found that a combination of different stresses, namely phosphate starvation, high pH, and excess nitrogen, triggers this response. These stresses can also be observed in C. crescentus’ natural freshwater habitat during algae blooms. Furthermore, our results indicate that filamentous cells are able to reach beyond biofilm surfaces, possibly enabling cells to reach nutrients and to release progeny.

While our studies highlight that cell filamentation is a common bacterial response to stress, some stress conditions, such as acute proteotoxic stress, lead to a growth arrest. In paper III we show that the regulatory interaction between the major chaperone DnaK and the heat shock sigma factor σ32 adjusts growth rate in response to changes of the global protein folding state. We show that high σ32 activity inhibits growth by re-allocating cellular resources from proliferative to maintenance functions. Under stress conditions when σ32 is active, this re-allocation likely helps cells to survive. However, under non-stress conditions unrepressed σ32 activity is detrimental. We demonstrate that in the absence of stress, the DnaK chaperone is absolutely necessary to limit σ32 activity and in this way to allow rapid proliferation.

In summary, the described studies highlight critical pathways that allow C. crescentus to integrate environmental information with cell cycle and growth regulation and shed new light onto the mechanisms by which bacteria adapt to their environment and in this way ensure their survival.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2018
Keywords
Caulobacter crescentus, bacteria, cell cycle, stress, filamentation, cell morphology, DNA replication, cell division
National Category
Microbiology
Research subject
Molecular Bioscience
Identifiers
urn:nbn:se:su:diva-158108 (URN)978-91-7797-381-2 (ISBN)978-91-7797-380-5 (ISBN)
Public defence
2018-09-21, Vivi Täckholmsalen, NPQ-huset, Svante Arrhenius väg 20 A, Stockholm, 10:00 (English)
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Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 2: Manuscript.

Available from: 2018-08-29 Created: 2018-07-27 Last updated: 2018-09-27Bibliographically approved
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